Abstract
The corticotropin-releasing factor (CRF) and its type 1 receptor (CRF(1)R) play a key role in the regulation of the hypothalamic-pituitary-adrenal (HPA) axis. Dysregulation of the HPA axis is associated with congenital adrenal hyperplasia (CAH) and depression. Non-peptide CRF(1)R-selective antagonists displayed antidepressant effects on animal models and are used for the management of CAH. To develop novel non-peptide CRF(1)R antagonists, we have previously designed and synthesized a series of substituted pyrimidines. Among these analogs, molecule 43 (M43) binds to CRF(1)R with the highest affinity. Based on this finding, we selected M43 for further pharmacological characterization in the present study. The results suggest that M43 is a potent CRF(1)R antagonist, blocking the ability of the CRF-related agonist, Tyr(0)-sauvagine, to stimulate (1) cAMP accumulation in HEK 293 cells expressing CRF(1)R and (2) the proliferation rate of RAW 264.7 macrophages. Computational studies suggest that the antagonist properties of M43 are mostly attributed to its ability to interact with residues in the allosteric pocket of CRF(1)R, comprised of the third, fifth, and sixth transmembrane domain residues, which block activation-associated structural rearrangements of the receptor. Our data will be used to design novel non-peptide CRF(1)R antagonists for clinical use.